The proposed research investigates the enzymology of influenza virus RNA replication; the influenza RNA-dependent RNA polymerase is a multi-subunit enzyme central to viral mRNA expression and viral RNA replication with no eukaryotic analogous cellular counterpart. Currently no influenza anti-viral drugs are aimed against this enzyme. Mechanisms involved in viral mRNA have been elucidated using reconstituted polymerase complexes expressed in the absence of influenza virus infection. These polymerase complexes do not efficiently catalyze viral RNA replication. To elucidate the mechanism of viral RNA replication and facilitate creation of new anti-viral therapies, we propose to utilize influenza polymerase complexes purified from the nuclei of influenza-infected cells. Specific aims are: 1) Establish requirements for virus RNA replication in vitro using purified influenza virus polymerase complexes from influenza virus infected cell nuclear extracts. 2) Identify modifications of polymerase subunits and host cellular factors that are essential for influenza virus RNA replication. 3) Determine template RNA requirements for vRNA replication by assembling initiation complexes that function in vRNA replication utilizing template-free polymerase complexes purified from influenza infected cell nuclear extracts and exogenous template RNAs. [unreadable] [unreadable]